N-glycosylation of the carcinoembryonic antigen related cell adhesion molecule, C-CAM, from rat liver: detection of oversialylated bi- and triantennary structures

Rat C-CAM is a ubiquitous, transmembrane and carcinoembryonic antigen relat ed cell adhesion molecule. The human counterpart is known as biliary glycop rotein (BGP) or CD66a. It is involved in different cellular functions rangi ng from intercellular adhesion, microbial receptor activity, signaling and tumor suppression. In the present study N-glycosylation of C-CAM immunopuri fied from rat liver was analyzed in detail. The primary sequence of rat C-C AM contains 15 potential N-glycosylation sites. The N-glycans were enzymati cally released from glycopeptides, fluorescently labeled with 2-aminobenzam ide, and separated by two-dimensional HPLC. Oligosaccharide structures were characterized by enzymatic sequencing and MALDI-TOF-MS. Mainly bi- and tri antennary complex structures were identified. The presence of type I and ty pe II chains in the antennae of these glycans results in heterogeneous glyc osylation of C-CAM. Sialylation of the sugars was found to be unusual; bi- and triantennary glycans contained three and four sialic acid residues, res pectively, and this linkage seemed to be restricted to the type I chain in the antennae. Approximately 20% of the detected sugars contain these unusua l numbers of sialic acids, C-CAM is the first transmembrane protein found t o be oversialylated.